Electron microscopy (EM) offers far better spatial resolution than fluorescence microscopy and therefore is a very important tool for cell biology. For example, mitochondria are just a few pixels wide by fluorescence but details and subcompartments can be seen by EM. However, a major limitation of EM is the lack of a fluorescent protein equivalent for highlighting specific proteins of interest.
Two existing genetically encoded reporters for EM are Horse Radish Peroxidase (HRP) and mini-singlet oxygen generator (SOG), both generating contrast by catalyzing the polymerization of a diaminobenzidine (DAB) into an osmiophilic polymer. Photo oxidation of miniSOG requires laser and blown oxygen. As such, use of miniSOG as an EM reporter is limited to small fields of view. HRP is a much easier to use, less temperamental, and more robust reporter than miniSOG, but it only works in the secretory pathway, such as in the endoplasmic reticulum (ER) and the Golgi apparatus, or on cell surfaces. It is inactive in any other cellular compartment, e.g., cytosol, due to disruption of the four disulfide bonds in this enzyme. Other reporters are prone to inactivation due to the strong fixation typically employed in EM.
It is of great importance to develop sensitive and robust reporters for use in EM.